Genetic profiling of drug resistance in Plasmodium falciparum

Abstract:

Malaria is an infectious febrile illness caused by four species of Plasmodium parasite, with Plasmodium falciparum responsible for the greatest number of fatalities. Unfortunately, drug resistant strains of P. falciparum are increasingly prevalent. Resistance to sulfadoxine-pyrimethamine (SP) is due to point mutations in the gene that encodes dihydrofolate reductase (dhfr). Patients infected with a parasite carrying three mutations in dhfr (N51I/C59R/S108N) are at elevated risk of failing SP treatment. Prior studies of the extended haplotype encompassing dhfr suggest that a single triple-mutant allele of dhfr emerged in Asia and spread to Africa. However, it is unclear whether this "Asian" strain replaced triple-mutants that had previously evolved in Africa, or simply invaded a population devoid of triple-mutants. In addition, prior studies were hampered by an inability to study infections containing DNA from multiple parasite strains. This dissertation presents a novel method for using yeast to separate haplotypes in mixed malaria infections. Second, it investigates the history of the Asian strain in Africa by analyzing samples collected in Kilifi, Kenya, from 1987-88 and 1993-95, periods immediately before and after the widespread use of SP. We genotyped each sample at dhfr and flanking microsatellite loci. All of the triple-mutants had the same haplotype, and it matched the haplotype of the Asian strain from previous studies. The wild-type parasites had a variety of haplotypes, none related to the triple-mutant haplotype. Each double-mutant (N51I/S108N or C59R/S108N) had a single haplotype. Both of the double-mutant haplotypes shared some alleles with wild-type samples and with each other, but neither shared any alleles with the triple-mutant haplotype. In addition, the Asian triple-mutant was present in 1988, well before the widespread use of SP in Kenya. These results indicate that the Asian triple-mutant arrived in Kenya before any SP use and rose to high frequency as soon as SP was introduced.